SU MMARY STATEMENT Anthrax toxin is the major cause of death in inhalation of Bacillus anthracis spores. It is composed of three proteins: protective antigen (PA), lethal factor (LF) and edema factor (EF). To express toxicity, PA binds to a cell surface receptor, tumor endothelial marker 8 (TEMS) or capillary morphogenesis gene-2 (CMG2). After proteolytic processing, the oligomerized PA binds to either LF or EF and transports the toxins to endosomes and cytosol. Within the cytosol, EF expresses its adenylyl cyclase activity and LF, a metalloprotease, hydrolyzes members of the mitogen-activated protein kinase (MAPK) kinase family. Therefore, interfering with the toxin functions in the above pathway is an important therapeutic approach. Here we propose a two-prong research. The first is to inactivate and remove LF in the blood plasma during the window of opportunity at an early stage of infection. As a metalloprotease, LF has a narrow substrate specificity and is not inactivated by alpha2-macroglobulin (alpha2M) that are present in the blood at high amounts. We propose to engineer the bait region of alpha2M to gain a new activity for LF inactivation. The new alpha2M may be infused into spore inhalation patients to keep them alive while being treated with antibiotics. The second approach is to interfere with the binding of PA to its cell surface anthrax toxin receptors. We propose to determine the three-dimensional structures of PA-TEM8 and PA-MG2 complexes and use the structural information to design small-molecular inhibitors that interfere with the complex formation. Various milestones are installed in the Research Plan and the final products are planned for human clinical experiments at the end of this program.